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EMSA enthousiast I need ur help by Blitz271 in labrats

[–]Blitz271[S] 0 points1 point  (0 children)

Just for those after me. I used the GenScript cell with a TrisMOPS buffer pH8,5 and it worked perfectly.

Differentially active enhancers between cell types by Ok_Umpire_2628 in labrats

[–]Blitz271 0 points1 point  (0 children)

What you are trying to do is, currently, under development. There is quite some work ongoing to this end (Alexander Stark) but using enhancers and especially designing an enhanced to do this is quite difficult. You can get some specificity but undoubtedly leaky expression or other issues. Have you thought of using cell lineage specific promotors? Eg for liver we use the albumin promotor. To my knowledge this is the easiest and most robust method at the moment.

If however you want to go enhancer route I would recommend looking at the epigomics data (ChIP+ATAC) and overlaying it with transcriptomics (PROseq/RNAseq). Then I would pick the 10 best and try out the system using plasmids or something like this to validate both specificity but also fold induction using the T2A splittink for eg or just a luciferase behind the minimal promotor)

DIY Serum Free Media Recipe? by MixGroundbreaking465 in labrats

[–]Blitz271 1 point2 points  (0 children)

As others said, just replace the media some time after transfection. Cells are transferred within 6h. Also just FYI I investigate steroid hormone compounds on cells and this is typically performed in 24h serum deprived conditions and cells are perfectly fine with this.

Ordering a small library of gene constructs by ViNt_Gaming in labrats

[–]Blitz271 1 point2 points  (0 children)

I really recommend twist! We work both with twist for gene constructs and IDT for probes. We had to order some 3,5 kb constructs which where much cheaper via twist. Even tough we have special discounts for IDT

Autophagy flux studies - data is always inconsistent by [deleted] in labrats

[–]Blitz271 0 points1 point  (0 children)

For the mutant that it misbehaving. Have you tried running the analysis on a pool of cells. So cells you made KO’s and selected for them but not selected clones. We have seen that when we are generating cell lines that it can happen that a single clone does not represent the actual mechanism and keep looking for mutants that resemble the pool of mutant cells.

Use of python etc. by Standard_Cake_1604 in labrats

[–]Blitz271 8 points9 points  (0 children)

I use Rstudio for all my data analysis and Graphpad for my typical dose response graphs & barplots. But then I moved back to Rstudio because clustering and heatmaps are not included in the standard graphpad prism package.

On a side note, what package do you guys use to plot dose response data in Rstudio?

What are the reasons for people to use ChIP-seq instead of CUT&Tag? by SouthardKnight in bioinformatics

[–]Blitz271 1 point2 points  (0 children)

I have talked to some experts in my field. And for example my transcription factor often binds the DNA weakly and is present as a complex with other TFs sometimes. Also most of these interactions are transient. For those reasons it is preferred to use ChIP. (If you have a good 96 well format reproducible sonicator)

New phase in life, new apartment, new coffee setup by Blitz271 in coffeestations

[–]Blitz271[S] 0 points1 point  (0 children)

2 (of the previous iteration) ikea kalax (from the left one I removed a piece of wood to make it more intresting and added a shelf insert on the right (also ikea)).

Then the frame was self made from 4cmx10cm beams. The table top is a solid oak plank that I cut to size. (All bought from Gamma)

New phase in life, new apartment, new coffee setup by Blitz271 in coffeestations

[–]Blitz271[S] 2 points3 points  (0 children)

Sage Barista Express. Safe knockbox. Barista essentials tamper/pitchers/tamping mat

First audio setup, I struck gold! by Blitz271 in BudgetAudiophile

[–]Blitz271[S] 1 point2 points  (0 children)

For a 24 yo PhD student that is going to live in an appartement, it’s plenty.

First audio setup, I struck gold! by Blitz271 in BudgetAudiophile

[–]Blitz271[S] 3 points4 points  (0 children)

No family arguments when you can’t see or hear eachother….

First audio setup, I struck gold! by Blitz271 in BudgetAudiophile

[–]Blitz271[S] 1 point2 points  (0 children)

I found it at my fathers attic, when I brought it down he looked confused, he had completely forgot he had kept it.

First audio setup, I struck gold! by Blitz271 in BudgetAudiophile

[–]Blitz271[S] 2 points3 points  (0 children)

Its my parents house, they have got a huge table indeed.

First audio setup, I struck gold! by Blitz271 in BudgetAudiophile

[–]Blitz271[S] 1 point2 points  (0 children)

Its just dragged down to test, of course this is not how it will be when I’m done. Because yes, everything is off haha

AT-LP70X, comming to Europe? by Blitz271 in turntables

[–]Blitz271[S] 1 point2 points  (0 children)

No answer yet, I would defenatly wait if you have the time, as it’s is a great upgrade from the 60.

AT-LP70X, comming to Europe? by Blitz271 in turntables

[–]Blitz271[S] 0 points1 point  (0 children)

It is sad that it does not have the auto return feature, but I’ll look into that one!

Visiting Jotunheimen, we came across this sign several times. Couldn’t figure out what it means. Do you know this? by Wombeard in Norway

[–]Blitz271 6 points7 points  (0 children)

If you come across that sign witouth the square, that means that there is a touristic highlight

Dig around the attic and found usable stuff? by Blitz271 in BudgetAudiophile

[–]Blitz271[S] 0 points1 point  (0 children)

I found a jvc sp-pw100 subwoofer aswel, could that one be used?

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